小球藻-大肠杆菌共培养产油过程控制研究及藻菌相互作用机制的解析

With the depletion of the nonrenewable petrochemical resources, biodiesel will be a promising alternative towards the petroleum fuel, while the production problem of microbial lipid has imposed restrictions on the development of the biodiesel industry. Although the algae and bacterium co-culture system can play important roles in developing the technology of microbial lipid production, the efficient theoretical guidance for the co-culture system is not available yet. For in depth understanding the co-culture system, we put forward to investigate the process control for the co-culture system in the present project, and then reveal microbial interactions through the methods of microbiome and quorum sensing analysis in the co-culture system. Firstly, the Chlorella vulgaris– Escherichia coli co-culture system for lipid production will be established. Secondly, the pH control strategy based on the carbon sources consumption and the flashing light effects will be investigated for the co-culture system. Thirdly, the C. vulgaris– E. coli interactions in the co-culture systems will be investigated to find the rules of material exchange and energy transfer, to obtain the key signal molecules and to analyze the mechanism of gene expression regulation through transcriptome analysis, proteome analysis, metabolome analysis and quorum sensing analysis. The mechanisms of C. vulgaris– E. coli interactions and the lipid production in the co-culture system will be put forward and verified, which could provide the scientific basis for the establishment of co-culture systems and rational design of microbial community. Besides, this study is also going to be an important case in point of microbiome.

随着化石资源的日益枯竭，微生物制备生物柴油将成为替代石化能源的重要方法。藻菌共培养体系对于实现高效稳定连续的油脂生产能够起到关键作用，但是藻菌共培养的研究还处在起步阶段，缺乏有效的理论指导。本项目提出对小球藻-大肠杆菌产油共培养体系进行过程控制研究，并采用微生物组学方法，通过转录组学、蛋白质组学、代谢物组学和群体效应分析，对小球藻和大肠杆菌的相互作用关系进行研究，获得物质交换和能量传递的规律，寻找关键的信号分子，解析基因表达调控机制，确定代谢途径的互补性和差异性，提出并验证微生物共生和油脂大量合成的机制。本研究将为高效稳定共培养方法的建立提供翔实的科学依据，为微生物群落的理性设计提供理论依据和现实例证，并将推动微生物组学的发展。

具有代谢互补特性的光自养与异养微生物进行共培养，可以实现微生物连续培养所需的微生态平衡。高效的连续培养通常具有更高的生产强度，更低的生产成本和需要更少投资的特点，这有利于生物燃料和生物化学品的商业化生产。然而，目前对微藻-细菌相互作用的研究还很有限，生物燃料和生物化学品的生产主要以分批或补料分批的方式进行。本项目建立了小球藻和大肠杆菌共培养体系，用于异戊二烯的生物合成。通过组学分析进一步探讨了共培养体系中微生物相互作用机制。小球藻促进了异戊二烯的合成和大肠杆菌的生长，而大肠杆菌则抑制了小球藻的生长。大肠杆菌与小球藻之间的相互作用与光合异养代谢产生的氧化压力密切相关。外源性葡萄糖的消耗导致过量的光电子，随后产生有毒的活性氧(ROS)。ROS可能导致蛋白质变性、DNA损伤和脂质过氧化等氧化压力。细胞内抗氧化和修复系统的活性反映了氧化压力。观察到小球藻对氧化压力的细胞响应是由Fenton反应触发的。小球藻可能通过种间交养的方式来保护大肠杆菌抵抗氧化压力，促进大肠杆菌的生长。在小球藻中，半胱氨酸的生物合成被大幅上调以减少ROS，同时小球藻可能为大肠杆菌提供了抗氧化所需的半胱氨酸。对小球藻和大肠杆菌共培养的研究对于揭示连续培养中光自养生物和异养生物之间的共同的相互作用具有重要意义。